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WO2025018275A1 - Composition contenant une bactérie lactique appartenant au genre lactococcus et son produit de fermentation - Google Patents

Composition contenant une bactérie lactique appartenant au genre lactococcus et son produit de fermentation Download PDF

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Publication number
WO2025018275A1
WO2025018275A1 PCT/JP2024/025193 JP2024025193W WO2025018275A1 WO 2025018275 A1 WO2025018275 A1 WO 2025018275A1 JP 2024025193 W JP2024025193 W JP 2024025193W WO 2025018275 A1 WO2025018275 A1 WO 2025018275A1
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Prior art keywords
bone
lactic acid
lactococcus
decrease
composition
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English (en)
Japanese (ja)
Inventor
諭 高杉
公一郎 角
恭子 伊藤
健太郎 中村
圭介 古市
麻美 土屋
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Meiji Co Ltd
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Meiji Co Ltd
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    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES, NOT OTHERWISE PROVIDED FOR; PREPARATION OR TREATMENT THEREOF
    • A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
    • A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
    • A23L33/135—Bacteria or derivatives thereof, e.g. probiotics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/185—Acids; Anhydrides, halides or salts thereof, e.g. sulfur acids, imidic, hydrazonic or hydroximic acids
    • A61K31/19—Carboxylic acids, e.g. valproic acid
    • A61K31/195—Carboxylic acids, e.g. valproic acid having an amino group
    • A61K31/197—Carboxylic acids, e.g. valproic acid having an amino group the amino and the carboxyl groups being attached to the same acyclic carbon chain, e.g. gamma-aminobutyric acid [GABA], beta-alanine, epsilon-aminocaproic acid or pantothenic acid
    • A61K31/198—Alpha-amino acids, e.g. alanine or edetic acid [EDTA]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
    • A61K35/66—Microorganisms or materials therefrom
    • A61K35/74—Bacteria
    • A61K35/741—Probiotics
    • A61K35/744—Lactic acid bacteria, e.g. enterococci, pediococci, lactococci, streptococci or leuconostocs
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P19/00—Drugs for skeletal disorders
    • A61P19/08—Drugs for skeletal disorders for bone diseases, e.g. rachitism, Paget's disease
    • A61P19/10—Drugs for skeletal disorders for bone diseases, e.g. rachitism, Paget's disease for osteoporosis

Definitions

  • Probiotics are also known to have a positive effect on bone metabolism, and the mechanisms of this include increased mineral solubility due to the short-chain fatty acids produced, cancellation of mineral absorption inhibition by phytic acid due to the produced phytase enzyme, and suppression of intestinal inflammation (Non-Patent Document 29).
  • Patent Document 1 describes a bone strengthening composition containing as an active ingredient a bacterial cell and/or a culture of a bacterium belonging to the genus Lactobacillus, Lactococcus, or Bifidobacterium.
  • This document shows the results of an experiment in which the proliferation of osteoblasts was promoted when freeze-dried bacterial cells of specific strains of Lactobacillus delbrueckii subsp. bulgaricus, Lactobacillus helveticus, Lactobacillus gasseri, Lactococcus lactis subsp. lactis, and Bifidobacterium breve were added to a culture system of an established osteoblast cell line (MC3T3-E1).
  • this document also describes that, based on the results of an investigation into the effects of administering the H61 strain to young to adult mice, the H61 strain does not affect bone formation, and that administration of killed cells of lactic acid bacteria other than H61 (other strains of cremoris and Lactococcus lactis subsp. Lactis G50) did not result in a reduction in aging scores.
  • Lactococcus lactis is a typical lactic acid bacterium used as a cheese starter.
  • Lactococcus lactis ssp. lactis strains, Lactococcus lactis ssp. cremoris strains, Lactobacillus reuteri strains, and Lactobacillus brevis strains are known as lactic acid bacteria with the metabolic ability to convert arginine in the culture medium to ornithine (Patent Documents 2 and 3).
  • CremorisC60 restores T Cell Population in Small Intestinal Lamina Propria in Aged Interleukin-18 Deficient Mice.Nutrients. 2020 Oct 27;12(11):3287. doi: 10.3390/nu12113287. Kosaka A, Yan H, Ohashi S, Gotoh Y, Sato A, Tsutsui H, Kaisho T, Toda T, Tsuji NM. Lactococcus lactis subsp. cremoris FC triggers IFN- ⁇ product tion from NK and T cells via IL-12 and IL-18. Int Immunopharmacol. 2012 Dec;14(4):729-33. doi: 10.1016/j.intimp.2012.10.007. Epub 2012 Oct 23.
  • Yamamoto K Hosogaya N, Inoue T, Jounai K, Tsuji R, Fujiwara D, Yanagihara K, Izumikawa K, Mukae H.B. Efficacy of Lactococcus lactis strain plasma (LC-Plasma) in easing symptoms in patients with mild COVID-19: protocol for an exploratory, multicentre, double-blinded, randomized controlled trial (PLATEAU study).MJ Open. 2022 Sep 14;12(9):e061172. doi: 10.1136/bmjopen-2022-061172. Fujii T, Fujitomo T, Tsuji R, Kubo R, Kato Y, Kanauchi O.
  • gks6 supplementation ameliorates bone loss in ovariectomized mice by promoting osteoblast differentiation and inhibiting osteoclast formation.
  • Lactobacillus helveticus (ATCC 27558) upregulates Runx2 and Bmp2 and modulates b one mineral density in ovariectomy-induced bone loss rats.
  • Preventing osteoporosis is a very important issue from the perspective of preventing the need for support or nursing care, improving quality of life, and extending healthy life expectancy.
  • active ingredients that are effective against osteoporosis caused by various risk factors or causes.
  • the present invention provides the following:
  • a composition for treating osteoporosis comprising at least one of any lactic acid bacteria belonging to the genus Lactococcus and any fermentation product of any lactic acid bacteria belonging to the genus Lactococcus.
  • a composition comprising at least one of any lactic acid bacteria belonging to the genus Lactococcus and any fermentation product of any lactic acid bacteria belonging to the genus Lactococcus, for any one selected from the group consisting of increasing bone strength, suppressing a decrease in bone strength, increasing bone density, suppressing a decrease in bone density, enhancing bone quality, suppressing a decrease in bone quality, enhancing bone structure, suppressing a decrease in bone structure, promoting bone formation, suppressing a decrease in bone formation, reducing bone resorption, and suppressing an increase in bone resorption.
  • the composition according to 1 or 2 wherein the composition is for any one selected from the group consisting of increasing bone strength, inhibiting a decrease in bone strength, strengthening bone structure, and inhibiting a decrease in bone structure.
  • the present invention includes a lactic acid bacterium belonging to the genus Lactococcus, and the lactic acid bacterium belonging to the genus Lactococcus is Lactococcus lactis subsp. lactis, Lactococcus lactis subsp. hordniae, or Lactococcus lactis subsp. comptae.
  • composition belonging to The composition according to claim 1 or 2, wherein the composition is for any one selected from the group consisting of increasing bone strength, inhibiting a decrease in bone strength, increasing bone density, inhibiting a decrease in bone density, strengthening bone structure, and inhibiting a decrease in bone structure.
  • the lactic acid bacterium belonging to the genus Lactococcus is a lactic acid bacterium belonging to Lactococcus lactis subsp. lactis.
  • composition described in any one of 1 to 5 comprising a lactic acid bacterium belonging to the genus Lactococcus, wherein the lactic acid bacterium belonging to the genus Lactococcus is Lactococcus lactis subsp. lactis OLS3445 (accession number NITE BP-3930) or OLS3301 (accession number NITE BP-432).
  • lactis OLS3445 accession number NITE BP-3930
  • OLS3301 accession number NITE BP-432
  • lactis OLS3445 (accession number NITE BP-3930) or OLS3301 (accession number NITE BP-432).
  • a composition described in any one of 1 to 10 comprising a fermentation product of any lactic acid bacterium belonging to the genus Lactococcus, wherein the fermentation product does not contain bacterial cells.
  • a composition comprising at least one fermentation product of any of the lactic acid bacteria belonging to the genus Lactococcus and any of the lactic acid bacteria belonging to the genus Lactococcus, for use in a method for treating osteoporosis.
  • a method or non-therapeutic method for treating osteoporosis comprising a step of administering to a subject a composition comprising at least one fermentation product of any of the lactic acid bacteria belonging to the genus Lactococcus and any of the lactic acid bacteria belonging to the genus Lactococcus.
  • Use or non-therapeutic use of a composition comprising at least one fermentation product of any of the lactic acid bacteria belonging to the genus Lactococcus and any of the lactic acid bacteria belonging to the genus Lactococcus for the treatment of osteoporosis.
  • a composition comprising at least one of any lactic acid bacteria belonging to the genus Lactococcus and any fermentation product of any lactic acid bacteria belonging to the genus Lactococcus for use in a method for any selected from the group consisting of increasing bone strength, suppressing a decrease in bone strength, increasing bone density, suppressing a decrease in bone density, enhancing bone quality, suppressing a decrease in bone quality, enhancing bone structure, suppressing a decrease in bone structure, promoting bone formation, suppressing a decrease in bone formation, suppressing a decrease in bone resorption, and suppressing an increase in bone resorption.
  • any lactic acid bacteria belonging to the genus Lactococcus and any fermentation product of any lactic acid bacteria belonging to the genus Lactococcus in the manufacture of a composition for any selected from the group consisting of increasing bone strength, suppressing a decrease in bone strength, increasing bone density, suppressing a decrease in bone density, enhancing bone quality, suppressing a decrease in bone quality, enhancing bone structure, suppressing a decrease in bone structure, promoting bone formation, suppressing a decrease in bone formation, suppressing a decrease in bone resorption, and suppressing an increase in bone resorption.
  • a composition comprising at least one of any of the lactic acid bacteria belonging to the genus Lactococcus and a fermentation product of any of the lactic acid bacteria belonging to the genus Lactococcus, for any of the following selected from the group consisting of increasing bone strength, suppressing a
  • a lactic acid bacterium belonging to the genus Lactococcus is used, and the lactic acid bacterium belonging to the genus Lactococcus is Lactococcus lactis subsp. lactis, Lactococcus lactis subsp. hordniae, or Lactococcus lactis subsp. cortae. It is a lactic acid bacterium belonging to 24.
  • lactis OLS3445 accession number NITE BP-3930
  • OLS3301 accession number NITE BP-432
  • a composition for treating osteoporosis comprising at least one of any lactic acid bacteria belonging to the genus Lactococcus and any fermentation product of any lactic acid bacteria belonging to the genus Lactococcus.
  • a composition comprising at least one of any lactic acid bacteria belonging to the genus Lactococcus and any fermentation product of any lactic acid bacteria belonging to the genus Lactococcus, for any one selected from the group consisting of increasing bone strength, suppressing a decrease in bone strength, increasing bone density, suppressing a decrease in bone density, enhancing bone quality, suppressing a decrease in bone quality, enhancing bone structure, suppressing a decrease in bone structure, promoting bone formation, suppressing a decrease in bone formation, reducing bone resorption, and suppressing an increase in bone resorption.
  • composition according to 1 or 2 comprising lactic acid bacteria belonging to the genus Lactococcus, wherein the lactic acid bacteria belonging to the genus Lactococcus are lactic acid bacteria belonging to Lactococcus lactis.
  • composition according to any one of 1 to 3 comprising lactic acid bacteria belonging to the genus Lactococcus, wherein the lactic acid bacteria belonging to the genus Lactococcus are lactic acid bacteria belonging to Lactococcus lactis subsp. lactis.
  • composition according to any one of 1 to 4 comprising a lactic acid bacterium belonging to the genus Lactococcus, wherein the lactic acid bacterium belonging to the genus Lactococcus is Lactococcus lactis subsp. lactis OLS3445 (accession number NITE BP-3930) or OLS3301 (accession number NITE BP-432).
  • lactis OLS3445 accession number NITE BP-3930
  • OLS3301 accession number NITE BP-432
  • a composition comprising at least one fermentation product of any of the lactic acid bacteria belonging to the genus Lactococcus and any of the lactic acid bacteria belonging to the genus Lactococcus, for use in a method for treating osteoporosis.
  • a method or non-therapeutic method for treating osteoporosis comprising a step of administering to a subject a composition comprising at least one fermentation product of any of the lactic acid bacteria belonging to the genus Lactococcus and any of the lactic acid bacteria belonging to the genus Lactococcus.
  • Use or non-therapeutic use of a composition comprising at least one fermentation product of any of the lactic acid bacteria belonging to the genus Lactococcus and any of the lactic acid bacteria belonging to the genus Lactococcus for the treatment of osteoporosis.
  • a composition comprising at least one of any lactic acid bacteria belonging to the genus Lactococcus and any fermentation product of any lactic acid bacteria belonging to the genus Lactococcus for use in a method for any selected from the group consisting of increasing bone strength, suppressing a decrease in bone strength, increasing bone density, suppressing a decrease in bone density, enhancing bone quality, suppressing a decrease in bone quality, enhancing bone structure, suppressing a decrease in bone structure, promoting bone formation, suppressing a decrease in bone formation, suppressing a decrease in bone resorption, and suppressing an increase in bone resorption.
  • any lactic acid bacteria belonging to the genus Lactococcus and any fermentation product of any lactic acid bacteria belonging to the genus Lactococcus in the manufacture of a composition for any selected from the group consisting of increasing bone strength, suppressing a decrease in bone strength, increasing bone density, suppressing a decrease in bone density, enhancing bone quality, suppressing a decrease in bone quality, enhancing bone structure, suppressing a decrease in bone structure, promoting bone formation, suppressing a decrease in bone formation, suppressing a decrease in bone resorption, and suppressing an increase in bone resorption.
  • a composition comprising at least one of any of the lactic acid bacteria belonging to the genus Lactococcus and a fermentation product of any of the lactic acid bacteria belonging to the genus Lactococcus, for any of the following selected from the group consisting of increasing bone strength, suppressing a
  • compositions use in production, therapeutic or non-therapeutic method, or use or non-therapeutic use according to any one of claims 11 to 14, wherein the composition comprises a lactic acid bacterium belonging to the genus Lactococcus, and the lactic acid bacterium belonging to the genus Lactococcus is Lactococcus lactis subsp. lactis OLS3445 (accession number NITE BP-3930) or OLS3301 (accession number NITE BP-432).
  • Lactic acid bacteria belonging to the genus Lactococcus can be used to treat osteoporosis.
  • lactic acid bacteria belonging to the genus Lactococcus any one selected from the group consisting of increased bone strength, suppression of a decrease in bone strength, increased bone density, suppression of a decrease in bone density, enhanced bone quality, suppression of a decrease in bone quality, enhanced bone structure, suppression of a decrease in bone structure, promotion of bone formation, suppression of a decrease in bone formation, reduction in bone resorption, and suppression of an increase in bone resorption can be achieved.
  • Bone strength (A) Polar moment of area, (B) Minimum moment of area mean ⁇ standard error, *: significant difference between normal and control diet groups; a, b: significant difference between different letters among the four EtOH intake groups Femoral bone mineral density mean ⁇ standard error, a, b, c: Different letters indicate significant differences among the four EtOH intake groups Femur longitudinal sections (from left: normal group, control diet group, unfermented food group, 3445 group, Orn group) Lumbar L2-4 bone mineral density mean ⁇ standard error, *: significant difference between OVX+ control group and sham group (p ⁇ 0.05), a, b, c: significant difference between different letters among the four EtOH intake groups (p ⁇ 0.05) Lumbar L2-4 bone mineral density mean ⁇ standard error, *: significant difference between OVX+ control group and sham group (p ⁇ 0.05), #: p ⁇ 0.1 Mean TRAP activity staining intensity per cell area ⁇ standard deviation (3 cases for medium without RANKL, 6 cases for fermentation supernatant added 2 groups); **:
  • the present invention relates to a composition that contains at least one of any lactic acid bacteria belonging to the genus Lactococcus and any fermentation product of any lactic acid bacteria belonging to the genus Lactococcus as an active ingredient.
  • any means that the type and number are optional.
  • Constaining as an active ingredient refers to the fact that the composition uses an effective amount to exert the intended function, that the ingredient is specified in the label as contributing to the intended purpose, etc. In foods with functional claims, the active ingredient is sometimes called a functional ingredient (an ingredient that contributes to a specific health purpose (excluding those related to reducing the risk of disease)).
  • the composition of the present embodiment contains, as an active ingredient, any lactic acid bacteria belonging to the genus Lactococcus (also referred to as Lactococcus lactic acid bacteria or Lactococcus bacteria).
  • the composition contains as an active ingredient a Lactococcus genus lactic acid bacterium belonging to any of the following species: Lactococcus lactis, Lactococcus chungangensis, Lactococcus garvieae, Lactococcus piscium, Lactococcus plantarum, Lactococcus raffinolactis, Lactococcus fujiensis, Lactococcus taiwanensis, Lactococcus formosensis, Lactococcus hircilactis, Lactococcus laudensis, Lactococcus allomyrinae, Lactococcus kimchi, Lactococcus laudensis, Lactococcus nasutitermitis, Lactococcus petauri, Lactococcus reticulitermitis, Lactococcus termiticola.
  • the composition contains as an active ingredient any lactic acid bacteria belonging to the Lactococcus lactis species, preferably any of the following subspecies: Lactococcus lactis subsp. lactis, Lactococcus lactis subsp. cremoris, Lactococcus lactis subsp. hordniae, Lactococcus lactis subsp. cortae.
  • the composition comprises as an active ingredient a lactic acid bacterium belonging to Lactococcus lactis subsp. lactis, Lactococcus lactis subsp. hordniae, or Lactococcus lactis subsp. gagtae.
  • the composition comprises as an active ingredient a lactic acid bacterium belonging to Lactococcus lactis subsp. lactis.
  • the composition contains as an active ingredient either Lactococcus lactis subsp. lactis OLS3445 or Lactococcus lactis subsp. lactis OLS3301, preferably Lactococcus lactis subsp. lactis OLS3345.
  • a strain taxonomically equivalent to a certain strain refers to, for example, any of the following: A strain belonging to the same genus as strain S, the entire sequence of its 16S rRNA gene or a characteristic part thereof (such as the V1 region, the V2 region, or the entire or part of the V1 and V2 regions, or a part including the V1 and V2 regions, etc.) having a sequence identity of 90% or more, preferably 95% or more, more preferably 98% or more, even more preferably 98.5% or more, even more preferably more than 98.7%, even more preferably 99% or more, even more preferably 100% with the sequence of strain S; A strain that has the same scientific properties (sometimes called mycological properties) as strain S.
  • sequence identity means the percentage of the number of matching bases shared between two sequences when the sequences are optimally aligned.
  • Analysis of base sequence identity can be performed using algorithms or programs well known to those of skill in the art (e.g., BLASTN, BLASTP, BLASTX, ClustalW). When using a program, parameters can be appropriately set by those of skill in the art, or the default parameters of each program may be used. Specific techniques for these analysis methods are also well known to those of skill in the art. Commercially available genetic information processing software may be used to calculate identity.
  • the composition of the present embodiment contains a fermentation product of lactic acid bacteria belonging to the genus Lactococcus as an active ingredient.
  • the fermentation product may be contained in any state in the composition as long as it can exert the intended effect.
  • the fermentation product may not contain lactic acid bacteria cells.
  • the fermentation product may be, for example, a culture supernatant.
  • the fermentation product may also contain ornithine.
  • composition which contains as an active ingredient a fermentation product of lactic acid bacteria belonging to the genus Lactococcus and preferably does not contain lactic acid bacteria cells, is particularly suitable for use for any of the following selected from the group consisting of enhancing bone quality, inhibiting the decline in bone quality, promoting bone formation, inhibiting the decline in bone formation, reducing bone resorption, and inhibiting the increase in bone resorption.
  • the Lactococcus lactic acid bacteria and their fermentation products contained in the composition can be produced by culturing.
  • the culture conditions are not particularly limited as long as the desired effect is achieved.
  • Lactococcus lactic acid bacteria and their fermented products can be produced by culturing lactic acid bacteria according to standard methods.
  • Various media can be used for the culture, such as a milk medium or a medium containing milk components, a semi-synthetic medium not containing these, or a synthetic medium.
  • the fermented product of lactic acid bacteria can be obtained by inoculating the above sterilized medium with Lactococcus lactic acid bacteria and culturing it at about 20°C to 45°C for 5 hours to several days. The temperature and period of the culture can be appropriately adjusted depending on the Lactococcus lactic acid bacteria strain used and the desired number of bacteria.
  • the product obtained by the culture can be used as Lactococcus lactic acid bacteria or its fermented product as it is, or it can be used as Lactococcus lactic acid bacteria after processing such as centrifugation, membrane concentration, washing, drying, freeze-drying, sterilization, etc., or it can be used as a fermented product not containing bacteria after processing such as removing the bacteria and, if necessary, removing insoluble components and separating them by membrane.
  • the bacterial cells may be in a living state (viable bacterial cells) or in a dead state (dead bacterial cells) as long as they are capable of exerting the intended effect.
  • Killed bacteria can be obtained by sterilizing lactic acid bacteria.
  • the sterilization is not particularly limited as long as it can produce the desired effect, and can be by heating, germicidal lamp (UV), ozone, chemicals, high osmotic pressure, etc.
  • the killed bacteria are preferably heat-treated killed bacteria obtained by heat-treating live bacteria.
  • the heat treatment for obtaining heat-treated killed bacteria is not particularly limited as long as it can produce the desired effect, and is carried out at a temperature and time sufficient to kill the lactic acid bacteria used.
  • Such conditions vary depending on the lactic acid bacteria used, but are, for example, 55°C or higher, preferably 60°C or higher, more preferably 65°C or higher, even more preferably 70°C or higher, and may be 80°C or higher, or 90°C or higher.
  • the upper limit of the heat treatment temperature can be set appropriately, for example, 121°C or lower, 100°C or lower, 90°C or lower, or 80°C or lower.
  • the heat treatment time can be 1 minute or more, 3 minutes or more, 10 minutes or more, 15 minutes or more, 30 minutes or more, or 45 minutes or more.
  • the upper limit of the heat treatment temperature can be, for example, 120 minutes or less, 100 minutes or less, 90 minutes or less, or 80 minutes or less.
  • the lactic acid bacteria to be contained in the composition can be prepared in the form of washed cells, concentrate, dried material, suspension, paste, gel, etc.
  • composition of this embodiment may be used for the treatment of osteoporosis.
  • osteoporosis refers to a disease characterized by low bone mass and abnormalities in the microstructure of bone tissue, leading to increased bone fragility and increased risk of fracture.
  • risk factors and causes of osteoporosis include aging, menopause, smoking, alcohol intake, lack of exercise, endocrine disorders (hyperparathyroidism, Cushing's syndrome, hyperthyroidism, hypogonadism, etc.), nutritional (post-gastrectomy, anorexia nervosa, malabsorption syndrome, vitamin C deficiency, excess vitamin A or D, etc.), drug use (steroids, anticonvulsants, warfarin, sex hormone reduction therapy drugs, SSRIs, methotrexate, heparin, etc.), immobility (systemic (bed rest, paraplegia, disuse syndrome, space travel, etc.), local (post-fracture, etc.), congenital (osteogenesis imperfecta, Marfan syndrome, etc.), diabetes, rheumato
  • the composition of the present embodiment may be used for any of the following: Increased bone strength, suppression of decline in bone strength, increased bone density, suppression of decline in bone density, enhanced bone quality, suppression of decline in bone quality, enhanced bone structure, suppression of decline in bone structure, promotion of bone formation, suppression of decline in bone formation, decrease in bone resorption, suppression of increase in bone resorption, maintenance of bone health, and maintenance of bone components.
  • bone quality refers to factors other than bone density that affect bone strength.
  • Bone structure is one of the bone quality indicators.
  • Bone strength is measured by polar moment of area (strength against torsion) and minimum moment of area (strength against bending), and the composition of one embodiment can be used to inhibit an increase or decrease in at least one of polar moment of area and minimum moment of area.
  • Bone structure includes cortical bone structure and cancellous bone structure, and cortical bone structure is measured by cortical bone volume (Ct.V), cortical bone percentage (Ct.V/Tt.V (Tt.V is total bone volume)), cortical bone width (Ct.Th), cortical bone circumference (Ps.Pm), and cortical bone inner circumference (Ec.Pm), and the composition of one embodiment can be used to inhibit an increase or decrease in any of the cortical bone volume, cortical bone percentage, and cortical bone width in particular.
  • the bone structure of cancellous bone is measured using indices of trabecular width (Tb.Th), trabecular number (Tb.N), trabecular space (Tb.Sp), bone pattern factor (TBPf), and trabecular shape (SMI), and the composition of one embodiment can be used to inhibit an increase or decrease in any of the following: trabecular number, trabecular space, bone pattern factor, and trabecular shape.
  • Tb.Th indices of trabecular width
  • Tb.N trabecular number
  • Tb.Sp trabecular space
  • TBPf bone pattern factor
  • SMI trabecular shape
  • the location or shape of the bone is not particularly limited, and bones include femur, lumbar vertebrae, pelvis, patella, tibia, fibula, tarsus, metatarsus, humerus, ulna, radius, carpal bones, ilium, sacrum, coccyx, pubis, ischium, clavicle, scapula, cervical vertebrae, thoracic vertebrae, sternum, ribs, spine (backbone), frontal bone, temporal bone, cheekbone, maxilla, and mandible, but the composition of one embodiment can be used in particular for at least one of the femur, tibia, and lumbar vertebrae, more specifically the femur and the lumbar vertebrae. Furthermore, the composition of one embodiment can be used in particular for the treatment (particularly for reducing the risk or preventing) of hip fractures, vertebral fractures, and distal radius fractures.
  • the composition is suitable for use for any one selected from the group consisting of increasing bone strength, inhibiting bone strength loss, enhancing bone structure, inhibiting bone structure loss.
  • the active ingredient of such a composition is not particularly limited as long as it is a lactic acid bacterium belonging to the genus Lactococcus, but is preferably a lactic acid bacterium belonging to Lactococcus lactis, more preferably a lactic acid bacterium belonging to Lactococcus lactis subsp. lactis, Lactococcus lactis subsp. hordniae, or Lactococcus lactis subsp. cortae, and even more preferably a lactic acid bacterium belonging to Lactococcus lactis subsp. lactis.
  • the composition is suitable for use for any one selected from the group consisting of increasing bone strength, inhibiting bone strength loss, increasing bone density, inhibiting bone density loss, enhancing bone structure, inhibiting bone structure loss.
  • the active ingredient of such a composition is a lactic acid bacterium belonging to Lactococcus lactis subsp. lactis, Lactococcus lactis subsp. hordniae, or Lactococcus lactis subsp. gagtae, more preferably a lactic acid bacterium belonging to Lactococcus lactis subsp. lactis.
  • Bone mass is maintained by a balance between bone resorption and bone formation. Bone mass decreases with age in both men and women, but in particular in women, the balance between bone resorption and bone formation is disrupted due to the decline in estrogen associated with menopause, resulting in rapid bone loss.
  • Bone strength can be evaluated by bone density and bone quality. Bone quality is indicated by any of the following: bone microstructure, rate of bone turnover, presence or absence of microfractures, and mineralization density.
  • bone strength refers to a combination of two factors: bone density and bone quality. Bone density accounts for 70% of bone strength, and the remaining 30% is explained by bone quality. Bone quality is determined by microstructure, bone turnover (the balance between bone formation and bone resorption), microfractures (microcracks), and calcification of bone tissue ("Guidelines for the Prevention and Treatment of Osteoporosis, 2015 Edition (http://www.josteo.com/ja/guideline/doc/15_1.pdf), NIH Consensus Development Panel on Osteoporosis Prevention, Diagnosis and Therapy, "Osteoporosis prevention, diagnosis and therapy," JAMA. 2001;285(6):785-795.).
  • a certain component can be used for the application of this embodiment can be evaluated, for example, by the following indexes: ⁇ Bone density testing (DXA method, ultrasound method, MD method, QCT method, pQCT method) ⁇ X-ray examination ⁇ Height measurement ⁇ Blood test ⁇ Urine test to measure bone formation markers, bone resorption markers, bone quality markers, mineral metabolism markers, vitamin D, vitamin K nutritional status, etc. ⁇ Bone strength test (CT / finite element analysis, etc.)
  • treatment includes reducing the risk of onset, delaying onset, prevention, treatment, and halting or delaying progression.
  • Treatment includes radical treatment (treatment to remove the cause) and symptomatic treatment (treatment to improve symptoms).
  • Actions for improvement or treatment include medical actions performed by doctors and nurses and midwives under the instructions of doctors, as well as non-therapeutic actions performed by persons other than doctors, such as pharmacists, nutritionists (including registered dietitians and sports nutritionists), public health nurses, midwives, nurses, clinical laboratory technicians, sports instructors, pharmaceutical manufacturers, pharmaceutical distributors, food manufacturers, food distributors, etc.
  • prevention or reduction of the risk of onset includes recommendations for the intake of specific foods and nutritional guidance (including nutritional guidance necessary for the medical treatment of injured or sick persons, and nutritional guidance for maintaining and promoting health).
  • the composition is suitable for administration to subjects for whom it is desirable or necessary to prevent osteoporosis, or subjects who are aware of such a condition; subjects for whom it is desirable or necessary to prevent any of the following selected from the group consisting of increasing bone strength, inhibiting a decrease in bone strength, increasing bone density, inhibiting a decrease in bone density, enhancing bone quality, inhibiting a decrease in bone quality, enhancing bone structure, inhibiting a decrease in bone structure, promoting bone formation, inhibiting a decrease in bone formation, inhibiting a decrease in bone resorption, inhibiting an increase in bone resorption, maintaining bone health, and maintaining bone components; subjects for whom it is desirable or necessary to prevent osteoporosis, or subjects who are aware of such a condition, based on any of the following selected from menopausal time, medical history, lifestyle (diet, exercise, sleep, smoking history, drinking history, etc.), medication history, genetic polymorphism, etc.; women aged 40 or older; subjects for whom the risk of fracture is high despite having
  • Such subjects may be subjects selected based on the results of bone density tests (DXA, ultrasound, MD, QCT, pQCT); X-rays; height measurements; blood and urine tests to measure bone formation markers, bone resorption markers, bone quality markers, mineral metabolism markers, vitamin D, vitamin K nutritional status, and bone strength tests (CT/finite element analysis).
  • DXA bone density tests
  • X-rays ultrasound, MD, QCT, pQCT
  • height measurements blood and urine tests to measure bone formation markers, bone resorption markers, bone quality markers, mineral metabolism markers, vitamin D, vitamin K nutritional status, and bone strength tests (CT/finite element analysis).
  • CT/finite element analysis bone strength tests
  • the age of the subject to which the composition is administered is not particularly limited.
  • the subject may be, for example, a newborn (within 28 days of birth); an infant (less than 1 year of age); a toddler (1-6 years of age); a child (7 years of age or older, but less than 15 years of age); an adult (15 years of age or older); or a person 65 years of age or older.
  • the subject is a middle-aged or elderly person, for example, a person 40 years of age or older, a person 50 years of age or older, a person 60 years of age or older, a person 65 years of age or older, a person 70 years of age or older, or a person 75 years of age or older.
  • composition of the present invention can be a food composition or a pharmaceutical composition.
  • the food and pharmaceutical products include not only those for humans, but also those for animals other than humans, unless otherwise specified.
  • the food includes general food, functional food, and nutritional composition, as well as therapeutic food (for the purpose of treatment. A doctor issues a meal prescription, and a nutritionist or the like prepares a menu based on the prescription.), dietary therapy food, ingredient-adjusted food, nursing care food, and food for medical support, unless otherwise specified.
  • the food includes not only solids, but also liquids, such as beverages, drinks, liquid foods, and soups, unless otherwise specified.
  • Functional food refers to food that can impart a specific functionality to a living body, and includes, for example, health functional foods including foods with specified health uses (including conditional FOSHU [specially designated health foods]), functional food, foods with nutritional functions, foods with special uses, dietary supplements, health supplements, supplements (for example, tablets, coated tablets, sugar-coated tablets, capsules, liquids, and other various dosage forms), and beauty foods (for example, diet foods).
  • health functional foods including foods with specified health uses (including conditional FOSHU [specially designated health foods]
  • functional food foods with nutritional functions, foods with special uses
  • dietary supplements for example, tablets, coated tablets, sugar-coated tablets, capsules, liquids, and other various dosage forms
  • beauty foods for example, diet foods.
  • the term "functional food” includes health foods to which a health claim based on the food standards of Codex Alimentarius (Joint FAO/WHO Food Standards Commission) is applied.
  • composition of the present invention may be administered orally, parenterally, for example, via a tube (gastrostomy, enterostomy), intranasally, or orally.
  • the composition can be administered to a subject repeatedly, and can be administered to a subject continuously for an extended period of time.
  • the period is not particularly limited, but in order to fully demonstrate the effect, it is preferable to administer the composition continuously for a relatively long period of time, for example, 3 days or more, 1 week or more, 2 weeks or more, 1 month or more, 3 months or more, 6 months or more, or 1 year or more.
  • the composition may be administered routinely, proactively, such as when there is a high risk, or when the need arises.
  • the composition may be administered with a meal, before, after, or between meals, or at the onset of the disease or condition that it is desired to ameliorate with the composition.
  • the content of Lactococcus lactic acid bacteria in the composition of the present invention may be any amount that can achieve the desired effect.
  • the content of Lactococcus lactic acid bacteria in the composition can be appropriately set taking into consideration various factors such as the age, weight, and symptoms of the subject to be ingested or administered.
  • One unit of the composition can be the amount ingested or administered per day or once (which may be administered or ingested once a day, or multiple times a day, for example, three times a day).
  • the amount of lactic acid bacteria bacterial count
  • it is a value determined by a standard method, unless otherwise specified.
  • the intake amount of Lactococcus lactic acid bacteria can be 1 x 10 8 cells/day or more, 4 x 10 8 cells/day or more, 7 x 10 8 cells/day or more, 1 x 10 9 cells/day or more, 4 x 10 9 cells/day or more, or 7 x 10 9 cells/day or more.
  • the intake amount of Lactococcus lactic acid bacteria is preferably 1 x 10 10 cells/day or more, more preferably 4 x 10 10 cells/day or more, even more preferably 7 x 10 10 cells/day or more, even more preferably 1 x 10 11 cells/day or more, even more preferably 4 x 10 11 cells/day or more, 7 x 10 11 cells/day or more, 1 x 10 12 cells/day or more, even more preferably 4 x 10 12 cells/day or more, or 7 x 10 12 cells/day or more.
  • the upper limit can be set as appropriate, and whatever the lower limit, the number may be, for example, 1 x 10 14 cells/day or less, 7 x 10 13 cells/day or less, or 1 x 10 13 cells/day or less.
  • the content of Lactococcus lactic acid bacteria per unit of the composition may be 1 x 10 7 bacteria, 4 x 10 7 bacteria or more, 7 x 10 7 bacteria or more, 1 x 10 8 bacteria or more, 4 x 10 8 bacteria or more, or 7 x 10 8 bacteria or more.
  • the content of Lactococcus lactic acid bacteria per unit of the composition is preferably 1 x 10 9 bacteria or more, more preferably 4 x 10 9 bacteria or more, even more preferably 7 x 10 9 bacteria or more, even more preferably 1 x 10 10 bacteria or more, even more preferably 2 x 10 10 bacteria or more, 4 x 10 10 bacteria or more, 7 x 10 10 bacteria or more, even more preferably 2 x 10 11 bacteria or more, 4 x 10 11 bacteria or more, 7 x 10 11 bacteria or more, 1 x 10 12 bacteria or more, or 4 x 10 12 bacteria or more.
  • the upper limit can be set appropriately, and whatever the lower limit, the number may be, for example, 1 x 10 or less, 7 x 10 or less, 1 x 10 or less, or 7 x 10 or less. Note that there are commercially available products that contain about 1 x 10 lactic acid bacteria of the genus Lactococcus per product.
  • the composition may contain other active ingredients or nutritional ingredients that are acceptable as foods or pharmaceuticals.
  • ingredients include lipids (e.g., milk fat, vegetable oils and fats, oils and fats containing medium-chain fatty acids), proteins (e.g., milk proteins, milk protein concentrates (MPC), whey protein concentrates (WPC), whey protein isolates (WPI), ⁇ -lactalbumin ( ⁇ -La), ⁇ -lactoglobulin ( ⁇ -Lg), heat-denatured whey proteins, and enzyme-treated whey proteins), amino acids (e.g., lysine, arginine, glycine, alanine, glutamic acid, leucine, isoleucine, valine), and lactobacilli other than Lactococcus genus or Lactococcus genus milk.
  • lipids e.g., milk fat, vegetable oils and fats, oils and fats containing medium-chain fatty acids
  • proteins e.g., milk proteins,
  • Carbohydrates other than the constituent sugars of acid bacteria (glucose, sucrose, fructose, maltose, trehalose, erythritol, maltitol, palatinose, xylitol, dextrin), electrolytes (e.g., sodium, potassium, calcium, magnesium, phosphorus), vitamins (e.g., vitamin A, vitamin B1, vitamin B2, vitamin B6, vitamin B12, vitamin C, vitamin D, vitamin E, vitamin K, biotin, folic acid, pantothenic acid, and nicotinic acids), minerals (e.g., copper, zinc, iron, cobalt, manganese, chromium, molybdenum), antibiotics, dietary fiber, etc.
  • electrolytes e.g., sodium, potassium, calcium, magnesium, phosphorus
  • vitamins e.g., vitamin A, vitamin B1, vitamin B2, vitamin B6, vitamin B12, vitamin C, vitamin D, vitamin E, vitamin K, biotin, foli
  • the composition may further contain additives that are acceptable for use as food or medicine.
  • additives include inert carriers (solid or liquid carriers), excipients, surfactants, binders, disintegrants, lubricants, solubilizers, suspending agents, coating agents, colorants, preservatives, buffers, pH adjusters, emulsifiers, stabilizers, sweeteners, antioxidants, flavors, acidulants, and natural products.
  • these include water, other aqueous solvents, pharma- ceutical acceptable organic solvents, collagen, polyvinyl alcohol, polyvinylpyrrolidone, carboxyvinyl polymers, sodium alginate, water-soluble dextran, water-soluble dextrin, sodium carboxymethyl starch, pectin, xanthan gum, gum arabic, casein, gelatin, agar, glycerin, propylene glycol, polyethylene glycol, petrolatum, paraffin, stearyl alcohol, stearic acid, human serum albumin, mannitol, sorbitol, lactose, sucralose, stevia, aspartame, acesulfame potassium, citric acid, lactic acid, malic acid, tartaric acid, phosphoric acid, acetic acid, fruit juice, vegetable juice, etc.
  • pharma- ceutical acceptable organic solvents collagen, polyvinyl alcohol, polyvinylpyrrolidone, carboxyvinyl poly
  • the food composition of the present invention may be prepared in any form, such as solid, liquid, mixture, suspension, powder, granule, paste, jelly, gel, capsule, etc.
  • the food composition of the present invention may be prepared in any form, such as dairy products, supplements, confectionery, beverages, drinks, seasonings, processed foods, side dishes, soups, etc.
  • composition of the present invention may be in the form of liquid food, semi-liquid food, jelly, gel, powder, prepared milk powder, prepared liquid milk, powdered milk/liquid milk for pregnant women and lactating women, fermented milk, bar, mousse, chocolate, biscuit, ice cream, fermented milk, lactic acid bacteria beverage, dairy beverage, milk beverage, soft drink, tablet, cheese, bread, biscuit, cracker, pizza crust, food for sick people, nutritional food, frozen food, processed food, etc., and may be in the form of granule, powder, paste, thick liquid, etc. for mixing with beverages or foods for administration.
  • the granules and powders may be in the form of cubes or sticks (packed in a single serving).
  • modified powdered milk as defined in the Ministerial Ordinance on Compositional Standards, etc. of Milk and Dairy Products (hereinafter referred to as the "Milk, etc. Ordinance"), refers to raw milk, cow's milk, special milk, raw buffalo milk, or foods produced using these as ingredients, processed or as the main ingredient, to which nutrients necessary for infants have been added, and made into a powder.
  • modified liquid milk as defined in the Milk, etc. Ordinance, refers to raw milk, cow's milk, special milk, raw buffalo milk, or foods produced using these as ingredients, processed or as the main ingredient, to which nutrients necessary for infants have been added, and made into a liquid.
  • Formulated milk powder or liquid milk (these are sometimes collectively called formulated milk powders.
  • formulated milk powder is sometimes used as an example, but the explanation also applies to formulated liquid milk.
  • Preferred examples of formulated milk powders are for infants, follow-up use, low birth weight infants, and children.
  • the pharmaceutical composition of the present invention can be in any dosage form suitable for oral administration, including solid preparations such as tablets, granules, powders, pills, and capsules, liquid preparations such as solutions, suspensions, and syrups, gels, and aerosols.
  • the stage of blending the Lactococcus lactic acid bacteria can be appropriately selected.
  • the stage of blending is not particularly limited as long as the characteristics of the Lactococcus lactic acid bacteria are not significantly impaired.
  • a composition can be produced as fermented milk containing a sufficient amount of Lactococcus lactic acid bacteria by mixing milk, a dairy product, and water, homogenizing and sterilizing as necessary, inoculating with Lactococcus lactic acid bacteria, and fermenting.
  • the milk and dairy product used as raw materials can be appropriately selected according to the Lactococcus lactic acid bacteria used in fermentation.
  • a composition can be produced by mixing milk, a dairy product, and water, homogenizing and sterilizing as necessary, inoculating with lactic acid bacteria required for fermentation (e.g., Lactobacillus delbrueckii ssp. bulgaricus and S. thermophilus), and adding Lactococcus lactic acid bacteria having the ability to control bacteria belonging to the Fusobacterium genus during or after fermentation.
  • the raw milk and dairy products include milk of animal origin.
  • animal milk are cow's milk, goat's milk, sheep's milk, etc.
  • the raw milk and dairy products may also include milk of plant origin.
  • the raw milk and dairy products may be made of animal origin and may not include milk of plant origin. Examples of plant milk are almond milk, oat milk, coconut milk, rice milk, and hemp milk.
  • composition of the present invention can be labeled with its intended use (application), and can also be labeled with a recommendation for administration to a specific subject.
  • composition of the present invention may be labeled with its intended use (use), and in one embodiment, it may be labeled with a recommendation that the composition be administered to a specific subject. Examples of subjects to be labeled are as described above.
  • the composition of the present invention may be labeled with the function of the composition or active ingredient, or a method of use based on that function. For example, it may be labeled with the following: it can treat osteoporosis, increase bone strength, inhibit decrease in bone strength, increase bone density, inhibit decrease in bone density, enhance bone quality, inhibit decrease in bone quality, enhance bone structure, inhibit decrease in bone structure, promote bone formation, inhibit decrease in bone formation, reduce bone resorption, inhibit increase in bone resorption, maintain bone health, and maintain bone components.
  • Display can be explicit or implicit.
  • An example of explicit display is a direct statement on a tangible object such as the product itself, packaging, container, label, tag, etc.
  • an example of implicit display includes advertising and promotional activities by place or means such as websites, storefronts, pamphlets, exhibitions, seminars such as media seminars, books, newspapers, magazines, television, radio, mail, email, and voice.
  • the recommendation to take the composition is displayed personally.
  • a display can be made using a document (whether written or electronic) addressed to the subject, the subject's tablet device, smartphone, or personal computer, the subject's SNS, or the like.
  • a display can be made together with the display of the results of any test or analysis of the subject's bone mass, bone density, bone formation markers, etc., more specifically bone density tests (DXA, ultrasound, MD, QCT, pQCT); X-rays; height measurements; blood and urine tests to measure bone formation markers, bone resorption markers, bone quality markers, mineral metabolism markers, vitamin D, vitamin K nutritional status, etc.; bone strength tests (CT/finite element analysis), etc.
  • DXA ultrasound, MD, QCT, pQCT
  • X-rays X-rays
  • height measurements blood and urine tests to measure bone formation markers, bone resorption markers, bone quality markers, mineral metabolism markers, vitamin D, vitamin K nutritional status, etc.
  • bone strength tests CT/finite element analysis
  • a method for reducing the risk of osteoporosis, etc. in a subject includes a step of having a subject ingest a composition containing bacteria belonging to the genus Lactococcus based on the subject's bone mass, bone density, bone formation markers, bone fragility, fracture risk, etc., more specifically bone density tests (DXA, ultrasound, MD, QCT, pQCT); X-rays; height measurements; blood and urine tests to measure bone formation markers, bone resorption markers, bone quality markers, mineral metabolism markers, vitamin D, vitamin K nutritional status, etc., and bone strength tests (CT/finite element analysis).
  • DXA ultrasound, MD, QCT, pQCT
  • X-rays X-rays
  • height measurements blood and urine tests to measure bone formation markers, bone resorption markers, bone quality markers, mineral metabolism markers, vitamin D, vitamin K nutritional status, etc.
  • bone strength tests CT/finite element analysis
  • a method for suggesting to a subject to ingest a composition, a method for supporting a subject's diet, or a method for supporting a subject's health includes a step of suggesting to a subject to ingest a composition containing bacteria belonging to the genus Lactococcus based on the subject's bone mass, bone density, bone formation markers, bone fragility, fracture risk, etc.
  • Example 1 [the purpose] Alcohol intake is known to have a negative effect on bones by suppressing bone formation (Turner, 2000, Alcohol Clin Exp Res, 24: 1693-1701., Kanis JA et al., 2005, Osteoporosis Int, 16: 737-742.). Therefore, we investigated the effect of intake of fermented products of Lactococcus lactis spp. lactis OLS3445 on bones using chronic alcohol intake mice as a model of bone formation suppression.
  • the L-arginine in the medium in the table below was replaced with a non-essential amino acid mixture*1 so that the nitrogen content was equivalent, and a medium powder was used without adding water.
  • *1 Equal molar mixture of L-proline, L-alanine, L-histidine, L-glycine, L-aspartic acid, and L-serine
  • Test period 8 weeks
  • Dissection After fasting for about 4 hours, the animals were sacrificed after total blood collection from the abdominal vena cava under pentobarbital anesthesia. The right femur was removed and stored in 70% ethanol in a refrigerator.
  • X-ray CT measurement The femur was subjected to 1mm-wide cross-sectional imaging using an X-ray CT (Latheta, ALOKA), and bone strength (polar moment of area, minimum moment of area) and bone density were calculated using LaTheta software (version 1.31). Polar moment of area and minimum moment of area indicate strength against torsion and strength against bending, respectively.
  • Bone quality analysis The femur was photographed using ⁇ CT (ScanXmate-L080; Comscantecno Co., Ltd., Yokohama, Japan) under the conditions listed in the table below. The images were analyzed using TRI/3D-BON software (Ratoc System Engineering Co., Ltd., Tokyo, Japan), and indices related to the bone structure of the cortical and cancellous bone were calculated as bone quality indices.
  • Body weight, food intake, and water intake No significant differences were observed between groups in body weight, water or alcohol intake, food intake, or energy intake.
  • Bone Strength The polar moment of area, which is one of the bone strength indexes, showed a significant decrease due to alcohol intake (Fig. 1-A). On the other hand, no significant effect of alcohol intake was observed on the minimum moment of area (Fig. 1-B). Furthermore, among the alcohol intake groups, the polar moment of area in the 3445 group was significantly higher than all other groups, and the minimum moment of area was significantly higher than the non-fermented group. Therefore, it was shown that intake of the fermented OLS3445 strain suppresses the decrease in bone strength during chronic alcohol intake. Furthermore, it was shown that this effect is a unique effect of the fermented OLS3445 strain that is not observed in non-fermented or Orn.
  • bone strength bone density + bone quality
  • Bone density Alcohol intake did not significantly affect bone mineral density (Figure 2). On the other hand, the bone mineral density of the 3445 group was significantly higher than that of the other groups among the alcohol intake groups. Furthermore, this effect was unique to the fermented OLS3445 strain and not found in the non-fermented or Orn strains.
  • Bone quality bone structure: Next, the bone structure of the cortical and trabecular bones was measured.
  • the cortical bone volume (Ct.V) of the control diet group was significantly lower than that of the normal group.
  • no significant effect of alcohol intake was observed on the bone structure index of the trabecular bone. Therefore, the decrease in bone strength due to alcohol intake observed in this study was thought to be due to a decrease in the bone structure of the cortical bone, which is one of the bone quality indexes.
  • cortical bone volume (Ct.V), cortical bone ratio (Ct.V/Tt.V), and cortical bone width (Ct.Th) of the 3445 group were significantly higher than those of the non-fermented and Orn groups, and it was confirmed that the ingestion of the OLS3445 fermented product improved the bone structure of the cortical bone (table above).
  • the 3445 group showed an increase in trabecular number (Tb.N) compared to the other three groups, a decrease in trabecular space (Tb.Sp) compared to the control and Orn groups, a decrease in TBPf compared to the non-fermented group, and a decrease in SMI compared to the control group (table above). Therefore, it was shown that the OLS3445 fermented product has an effect of improving the bone structure of cortical and cancellous bone, which are bone quality indicators, and that this effect is unique to the OLS3445 fermented product. Furthermore, it was confirmed in the ⁇ CT images that the mesh structure of the cancellous bone of the 3445 group was significantly increased compared to the other groups ( Figure 3).
  • Example 2 [the purpose] We investigated the effect of fermented OLS3445 on bone metabolism using ovariectomized (OVX) rats with postmenopausal osteoporosis. It is known that bone resorption increases significantly during OVX. Therefore, the OVX rat model of postmenopausal osteoporosis is a model system in which increased bone resorption has a negative effect on bones.
  • OVX ovariectomized
  • Grouping The groups were divided so that the average values of body weight and bone density (measured by X-ray CT) were uniform among the groups. Since the fermentation product of OLS3445 contains Orn, a group was also provided with a diet containing the same amount of Orn.
  • (1) Sham group (2) OVX + control group (Cont group) (3) OVX + unfermented group (unfermented group) (4) OVX + OLS3445 strain fermentation product group (3445 group) (5) OVX + Orn group (Orn group) Eight animals with significant weight deviations the day after surgery were excluded.
  • Test period 8 weeks
  • Bone density measurement by X-ray CT Before surgery and at 4 and 8 weeks after the start of the study, X-ray CT (Latheta, Aloka) was used to take cross-sectional images of the second to fourth lumbar vertebrae (L2-4) at 1 mm intervals, and bone density was calculated.
  • the OLS3445 fermentation group showed a significantly higher bone mineral density than the Orn group at 4 weeks of intake, and compared with the control and Orn groups at 8 weeks of intake (Figure 4).
  • the OLS3445 fermentation group showed a tendency for higher cancellous bone mineral density at 4 and 8 weeks of intake compared with the control group ( Figure 5).
  • Example 3 [the purpose] We investigated the effects of ingestion of fermented Lc. lactis OLS3445 on bone formation markers in postmenopausal women.
  • the OLS3445 fermentation product was obtained in the same manner as in Example 1.
  • the OLS3445 fermentation product was freeze-dried to obtain a powder of OLS3445 heat-fermented bacteria fermentation product.
  • the study foods were blended and prepared as shown in the table below. The study foods were each packed in aluminum packages at 8 g per package, and subjects were asked to ingest 2 packages (16 g) per day.
  • Test item (1) Dextrin powder (2) OLS3445 heat-fermented bacteria powder (2 packets (16 g) contain 4 x 10 10 bacteria) (3) OLS3445 Heat-fermented bacteria powder (2 packets (16 g) contain 4 x 10 11 bacteria)
  • Selection criteria 1. Women who are 50 years of age or older at the time of obtaining consent, and who have been postmenopausal for 1 to 10 years. 2. Those with a BMI of 18.5 or greater and less than 30. 3. Those who have been fully informed of the purpose and contents of this study, are capable of consenting, fully understand the study, and volunteer and agree to participate of their own free will.
  • Exclusion Criteria 1. Those with food allergies, 2. Those with lactose intolerance, 3. Those who reached menopause before the age of 43, 4. Those who have been diagnosed with osteoporosis, 5. Those who have undergone bilateral oophorectomy, 6. Those who suffer from severe liver disorder, kidney or heart disease, organ disorders such as lungs or digestive organs (including gastrectomy), diabetes, rheumatoid arthritis, neurological or psychiatric disorders, endocrine disorders, or other severe or progressive diseases, 7. Those who have been diagnosed with congenital bone metabolic diseases (osteogenesis imperfecta, Marfan syndrome, etc.), 8. Those with a history of fracture within the past year, 9.
  • ⁇ Bone formation marker Serum BAP (bone alkaline phosphatase, CLEIA method)
  • Example 4 The inhibitory effect of the fermentation supernatant of Lc. lactis OLS3445 on osteoclast differentiation was examined using mouse macrophage-like cell line RAW264.7.
  • Cell experiments The procedure was as follows. (1) Grow the required number of RAW264.7 cells in standard growth medium (GM, DMEM, 10% FBS (inactivated), 1% penicilin-streptmycin). (2) Collect the cells, resuspend them in test growth medium (TM, ⁇ -MEM (containing no Phenol Red), 10% FBS (immobilized), 1% penicilin-streptmycin), and seed them into a 48-well plate at 1.5x10 ⁇ 4 cells/well and incubate at 37°C for 3-4 hours.
  • GM standard growth medium
  • FBS inactivated
  • TM test growth medium
  • TM was removed and replaced with osteoclast differentiation medium (DM; TM supplemented with 50 ng/mL sRANKL (Soluble RANK Ligand, Wako 184-01791)).
  • DM osteoclast differentiation medium
  • sRANKL Soluble RANK Ligand, Wako 184-01791
  • Fluorescence images were taken using a Keyence all-in-one fluorescence microscope BZ-X810 with Lysosensor filters (excitation 360/40, dichroic 400, fluorescence 540/40), GFP filter, and Cy5 filter, respectively. Images were captured in a 3x3 field of view using a 10x objective lens and linked to create a single image using the attached image analysis software.
  • Mature osteoclasts have high TRAP activity and are giant, multinucleated cells. For this reason, the standard method for evaluating osteoclast formation is to measure the number and area of TRAP-positive cells containing 3-5 or more nuclei (areas stained with F-actin) and the intensity of TRAP activity. However, this requires visual evaluation and is time-consuming. Therefore, in this study, we simply analyzed and quantified the TRAP staining intensity in the areas stained with F-actin, that is, the TRAP activity intensity per cell area, using the image analysis software Image-J (Rasband, W.S., ImageJ, U.S. National Institutes of Health, Bethesda, Maryland, USA, http://rsb.info.nih.gov/ij/, 1997-2012.), and used this as an index of osteoclast formation.
  • Image-J Rasband, W.S., ImageJ, U.S. National Institutes of Health, Bethesda, Maryland, USA, http://rsb
  • any of the lactic acid bacteria belonging to the genus Lactococcus or its fermented product is effective in treating osteoporosis caused by any risk factor and cause, and is also effective in reducing bone formation (including alcohol intake, aging, diabetes, renal failure, etc.) and increasing bone resorption (including menopause or ovariectomy, reduced female hormones, etc.).
  • Any lactic acid bacteria belonging to the genus Lactococcus or a fermentation product thereof can be used for increasing bone strength, suppressing a decrease in bone strength, increasing bone density, suppressing a decrease in bone density, enhancing bone quality, suppressing a decrease in bone quality, enhancing bone structure, suppressing a decrease in bone structure, promoting bone formation, suppressing a decrease in bone formation, reducing bone resorption, suppressing an increase in bone resorption, maintaining bone health, and maintaining bone components.
  • the present invention can provide a food composition and a method for producing a food that contribute to the treatment of osteoporosis, which is a widespread issue both domestically and internationally.
  • the present invention can also provide a food composition and a method for producing a food that supports the maintenance and improvement of people's health and the efficiency of work.
  • the present invention can support the maintenance and improvement of people's health and the efficiency of work.
  • the present invention can improve the nutrition of a variety of people, ensure healthy lives, and promote welfare.

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Abstract

La présente invention aborde le problème de la fourniture d'un moyen qui est efficace contre l'ostéoporose provoquée par diverses causes. L'invention concerne : une composition qui est destinée au traitement de l'ostéoporose et qui comprend l'une quelconque des bactéries lactiques appartenant au genre Lactococcus et/ou un produit de fermentation de l'une quelconque des bactéries lactiques appartenant au genre Lactococcus; et une composition pour une utilisation quelconque choisie dans le groupe consistant en la hausse de la résistance osseuse, l'inhibition d'une diminution de la résistance osseuse, l'augmentation de la densité osseuse, l'inhibition d'une diminution de la densité osseuse, l'amélioration de la qualité osseuse, l'inhibition d'une diminution de la qualité osseuse, l'amélioration de la structure osseuse, l'inhibition d'une diminution de la résistance osseuse, la promotion de la formation osseuse, l'inhibition d'une diminution de la formation osseuse, la diminution de la résorption osseuse, et l'inhibition d'une augmentation de la résorption osseuse. Les bactéries lactiques appartenant au genre Lactococcus appartiennent de préférence à Lactococcus lactis subsp. Lactis.
PCT/JP2024/025193 2023-07-14 2024-07-12 Composition contenant une bactérie lactique appartenant au genre lactococcus et son produit de fermentation Pending WO2025018275A1 (fr)

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Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2009066537A1 (fr) * 2007-11-19 2009-05-28 Meiji Dairies Corporation Inducteur de fonction immunorégulatrice et composition alimentaire
JP2017221157A (ja) * 2016-06-16 2017-12-21 株式会社明治 乳酸菌発酵促進剤
JP2018153177A (ja) * 2017-03-17 2018-10-04 島根県 桑実に由来する乳酸球菌、及びそれを含む経口組成物
WO2021112041A1 (fr) * 2019-12-02 2021-06-10 雪印メグミルク株式会社 Agent de renforcement osseux

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2009066537A1 (fr) * 2007-11-19 2009-05-28 Meiji Dairies Corporation Inducteur de fonction immunorégulatrice et composition alimentaire
JP2017221157A (ja) * 2016-06-16 2017-12-21 株式会社明治 乳酸菌発酵促進剤
JP2018153177A (ja) * 2017-03-17 2018-10-04 島根県 桑実に由来する乳酸球菌、及びそれを含む経口組成物
WO2021112041A1 (fr) * 2019-12-02 2021-06-10 雪印メグミルク株式会社 Agent de renforcement osseux

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